Authors:Ritu Gaur, Syed Imteyaz Alam, Dev Vrat Kamboj*

Title:Immunoproteomic analysis of antibody response of rabbit host against heat-killed Francisella tularensis Live Vaccine Strain

Journal nameCurrent Microbiology

Running title:Immunoproteomic analysis of Francisella tularensis

Authors:Ritu Gaur, Syed Imteyaz Alam, Dev Vrat Kamboj*

Affiliations: Biotechnology Division, Defence Research and Development Establishment, Gwalior-474002, India

*Corresponding

author: Biotechnology Division, Defence Research & Development Establishment, Gwalior-474002, India

Tel No. +917512347545, +917512390270

Fax No. +917512341148

Supplementary Table 1: Proteins identified from extractable proteome of Live Vaccine Strain of Francisella tularensis.

Spot ID / Protein (accession no.)a / Mowse score / Cov.b (%) / Mr/pIc / Mr/pId / Peptides (MS/MS) / Ion score / RMS Error
(ppm)
1 / Peptidoglycan-associated lipoprotein (gi|89255762) / 110 / 10 / 23.28/5.18 / 20.27/5.1 / TADYLIAHPDQPIK.
YNFNLGQK / 69
41 / 83
2 / Intracellular growth locus, subunit C (gi|89255550) / 836 / 53 / 22.12/5.65 / 20.41/5.9 / QQVTSGETIHVR
TDPTACIGSHPNCR
LFIDSLTIAGEK
LFIDSLTIAGEKLDK
NIVAIEGGEDVTKADSATAAASVIR
ADSATAAASVIR
AFSISIEPTELMGVSK
AFSISIEPTELMGVSK (M)
DGMSYHIISIDGLTTSQGSLPVCCAASTDK / 95
80
100
60
177
55
122
76
148 / 10
3 / Acetyl-CoA carboxylase, biotin carboxyl carrier protein subunit (gi|89256869) / 487 / 51 / 16.39/5.12 / 16.52/5.1 / VAEILNSSDIK
NNTAAITSVVSAAPVASNVASAAPAVATAATSAAAPK
VNLAEEISGEEIK
SPMVGTFYGASSPDAAPYVK (M) / 61
189
97
104 / 119
4 / FTL_1225 (Hypothetical protein) (gi|89256546) / 690 / 52 / 25.68/7.71 / 25.37/5.95 / SDSILVQVTGYATTTLEK
NIEQTTSNSGAINITVK
ISQSDLNNLQR
AFENQNK
LVVDVLDYNPPAR
DYLANFNK
QTNSNYTIQSIQYIDVDNYQPR
TAYSGDANSNSSNPVAVTQDINIK / 79
109
67
17
82
45
151
90 / 112
5 / ABC transporter, ATP-binding protein (gi|89256548) / 283 / 41 / 27.44/5.80 / 25.37/6.52 / DLNDLSISER
AAAGIFLSLQYPIEIPGVSNVQFLK
YMSRGVNEGFSGGEK
LAILDETDSGLDIDALQVVSQGANSMR (M)
SFLVITHYQR
LLDHIQPDFVHVLADGK / 32
22
-
27
68
104 / 110
6 / Rhodanese (gi|89256194) / 650 / 50 / 27.85/6.23 / 27.29/7.0 / MNETQTLDGLLIDTR
MNETQTLDGLLIDTR (M)
EESEVANGYIPNAIHLSK (NQ)
MYFYCGGGFR (C)
MYFYCGGGFR (C, M)
NVISVDGGWR
GYPTVSPNQFRPNEFLK
INSQELDGIVFDVR
EDSEFNR
IYLYCGSGFR (C)
SALAAESLQHMGYTNVVSIAGGIK (M) / 60
125
56
32
51
48
61
128
45
67
70 / 128
7 / Glutamate dehydrogenase (gi|89255702) / 893 / 49 / 49.13/6.49 / 49.44/7.4 / EVFSTLKPALEHNPK
YIEENILAR
VPWVDKDGNIQVNR
YQFNGAIGPFK (NQ)
FLGFEQVFK
EIGYMYGQYR
EIGYMYGQYR (M)
SLESGGSLIRPEATGYGAVFFLDEMLK (M)
GFVHDPEGITTDEKIEFLLK
EFNASWHAGQKPWGIK
ADIAIPAATQNEIDFEDAQK
YVVEASNMPTTNEAIEFLMK (2M)
AANAGGVAVSGLEMSQNSAR
LQQIMANIFHACK (C, M)
YNLGYNLVAGANLAGFEK / 24
51
113
56
66
37
19
50
82
59
97
7
-
36
127 / 127
8 / Inosine-5′-monophosphate dehydrogenase (gi|89256767) / 1060 / 49 / 52.06/7.19 / 56.79/7.75 / ITQQAITFDDVLLSPR
YSNVLPHQVDLK
DIQLNIPLVSAAMDTVTESR
DIQLNIPLVSAAMDTVTESR (M)
LAIAIAQEGGIGIIHK
EHNFSGFPVVDDNNK
DLDEPVSSIMTPR (M)
VAALAAEGVDIIVVDTAHGHSQGVLDTVK
DNYPHIQVIGGNIATAEAAK
IVAGVGVPQITAIANVAEALK
GTGVPVIADGGIR
AIVAGASVVMIGGLFAGTEESPGEVELFQGR (M)
KFVPEGVEGR
FVPEGVEGR
TEPTFVQITGAGFNESHVHNVTITK / 117
34
26
64
71
75
5
95
126
104
50
36
34
50
107 / 131
9 / Inosine-5′-monophosphate dehydrogenase (gi|89256767) / 995 / 40 / 52.06/7.19 / 57.17/7.9 / ITQQAITFDDVLLSPR
YSNVLPHQVDLK
DIQLNIPLVSAAMDTVTESR (M)
LAIAIAQEGGIGIIHK
RFENGMVIDPITIK (NQ, M)
EHNFSGFPVVDDNNK
DLDEPVSSIMTPR (M)
DNYPHIQVIGGNIATAEAAK
IVAGVGVPQITAIANVAEALK
GTGVPVIADGGIR
KFVPEGVEGR
FVPEGVEGR
TEPTFVQITGAGFNESHVHNVTITK / 131
65
40
86
2
71
13
150
92
38
40
54
142 / 132
10 / Glutamate dehydrogenase (gi|89255702) / 648 / 33 / 49.13/6.49 / 49.44/7.1 / YIDSVIAQVEK
EVFSTLKPALEHNPK
YIEENILAR
VPWVDKDGNIQVNR
FHPSVYSGIIK
FLGFEQVF
EIGYMYGQYR
EIGYMYGQYR (M)
GFVHDPEGITTDEKIEFLLK
ADIAIPAATQNEIDFEDAQK
LQQIMANIFHACK (C, M)
YNLGYNLVAGANLAGFEK / 18
25
52
54
63
59
28
20
17
117
22
135 / 98
11 / Molecular chaperone DnaK (gi|89256512) / 1378 / 33 / 69.14/4.88 / 73.47/5.0 / TTPSVVAYTDSGEILVGQAAK
RQAVTNPDNTFFAIK
QAVTNPDNTFFAIK
MAPPQVSAEVLR (M)
KTAEDYLGEPVTEAVITVPAYFNDSQR
TAEDYLGEPVTEAVITVPAYFNDSQR
IINEPTAAALAYGVDSK
EQGIDLHNDKLALQR
AKFESLVSDLVMR (M)
FESLVSDLVMR
SDITEVLLVGGQTR
KDVNPDEAVAVGAAIQGGVLAGDVK
KSQVFSTAEDNQPAVTIHVLQGER
SQVFSTAEDNQPAVTIHVLQGER
FDLADIPPAPR
FHDLVTAR
ALEEAFAPIAQK / 150
45
89
31
115
128
103
44
24
33
90
135
95
114
69
63
54 / 99
12 / Chaperonin GroEL (gi|89256983) / 560 / 18 / 57.37 /4.99 / 60.27/5.2 / QVLFSDEAR
TADVAGDGTTTATVLAQALLTEGLK
GFEDELDVVEGMQFDR
GFEDELDVVEGMQFDR (M)
ANIAEANSDYDREK
AAVEEGIVAGGGVALIR
ALDGLTGENDDQNHGIALLR / 45
116
139
62
39
114
106 / 108
13 / Chitinase (gi|89256806) / 429 / 11 / 83.54/4.96 / 80.06/5.1 / ADLVNFETTQLSNGEYLTK (NQ)
TLDASADSWGLSASAR
YYLAIAVNGAR (NQ)
KLIVGLPLYAR
LIVGLPLYAR
TFVTFDDVWSVNEK
YTIEVIASTSNASR / 56
72
51
62
65
66
58 / 101
14 / Peroxidase/catalase (gi|89256792) / 417 / 11 / 81.18/5.32 / 74.45/5.75 / YTQEFYNNPEEFKEEFAK
QVSTQDIAQLEQDIINSGLTNQQLIK
VSLADLIVLGGNVGVEQAAK
WTASPVDLIFGSNSELK / 80
121
145
71 / 109
15 / Malonyl CoA-acyl carrier protein transacylase (gi|167010384) / 611 / 31 / 33.48/5.24 / 33.93/5.4 / TAVVFPGQGSQK
LGMLQDYYENFETFR
LGMLQDYYENFETFR (M)
TEFTQPALLATSYAIYEVLK
DALQLVSTR
SCQEASDAGIVEAANFNSTGQVVISGEK (C)
LGVTEVIECGPNK (C) / 77
31
87
91
72
191
97 / 115
16 / Outer membrane protein FopA (gi|89256632) / 612 / 26 / 41.23/5.59 / 41.91/5.4 / YFAVQYNQLVGR
NVQASVDYR
YIQTMAPSNISGANGR (NQ)
YIQTMAPSNISGANGR (NQ, M)
AGTNMIGAGLTWFFGGK (M)
YVLPAGIK
AIASFVNFIK
GYASQGQTGSEFDIYNQK
GFGYNDTLGGIHK / 92
37
55
26
68
30
75
150
106 / 104
17 / Elongation factor Tu (gi|89257020) / 853 / 41 / 43.36/5.12 / 48.79/5.52 / GITINTSHVEYESPNR
CDMVDDEELLELVEMEVR (C, 2M)
ELLDQYEFPGDDTPVIMGSALR
ELLDQYEFPGDDTPVIMGSALR (M)
IVELVQAMDDYIPAPER
IVELVQAMDDYIPAPER (M)
DTEKPFILPIEDVFSISGR
GVVNVGDEVEVVGIRPTQK
LLDRGEAGDNVGILVR
GEAGDNVGILVR
FEAEVYVLSK
GYRPQFYFR
MTITLINPIAMDEGLR
MTITLINPIAMDEGLR (M) / 67
35
82
33
91
79
118
111
61
71
68
24
55
32 / 107
18 / Outer membrane protein FopA (gi|254369500) / 276 / 12 / 39.47/5.17 / 35.07/5.68 / YFAVQYNQLVGR
YIQTMAPSNISGANGR (NQ, M)
GYASQGQTGSEFDIYNQK / 87
18
150 / 118
19 / Outer membrane protein FopA (gi|254369500) / 508 / 20 / 39.47/5.17 / 34.38/6.1 / YFAVQYNQLVGR
NVQASVDYR
YIQTMAPSNISGANGR
YIQTMAPSNISGANGR (NQ. M)
AIASFVNFIK
GYASQGQTGSEFDIYNQK
GFGYNDTLGGIHK / 87
40
11
35
69
150
88 / 96
21 / Succinyl-CoA synthetase, alpha subunit (gi|89256838) / 230 / 18 / 30.10/6.24 / 44.48/6.9 / NGTFHSEQAIAYGTNIVGGVTPGK (NQ)
LVVIITEGVPTLDMLVVK (M)
FAAFEAAGIAYTR / 95
52
84 / 102
22 / Aspartyl/glutamyl-tRNA amidotransferase subunit A (gi|89257102) / 301 / 13 / 52.36/5.41 / 55.31/5.95 / VPGGSSGGSAAAVAAGFAPISTGSDTGGSVR
EALSTYYIITPAEAAANLAR
HYGIEEFILQQAR / 99
118
84 / 80
23 / Hypothetical protein FTL_0572 (gi|89255970) / 893 / 35 / 51.95/6.69 / 56.42/6.2 / SSAEEILNDVMENYIDQNNLR (M)
DRYDYVGSAIGTASVNQTNSNYVDSAQLAFEK
AQAEYISFISANTK
SLNIDSTQGSTANEIDTYADKPK
EQGLNPNDFATPEEK
SLTTGFGNLSGLLPIK
SGEDMMGDYGIR (2M)
RIDATDIEK
YKLPGTENIIYGVVLK
LPGTENIIYGVVLK
NFSYDEYR
NFSYDEYRK / 62
19
90
85
84
140
20
11
88
106
55
70 / 100
24 / Cell division protein FtsZ (gi|56707353) / 892 / 51 / 39.72/4.76 / 42.19/5.05 / MFDFNDSMVSNAIIK (2M)
VQNILQIGTNLTK
GLGAGANPEIGKR
EMGILTVAVVTKPFPFEGPR
EMGILTVAVVTKPFPFEGPR (M)
HVDSIITVPNEK
GASLIDAFNAANDVLGNAVK
GVSELITKPGLINVDFADVR
AVMTNMGLAMMGMGEASGENR (5M)
EAAEAAISSPLLEDINLDGAK
GVIVNITAGMDMSIGEFEEVGEVIR (2M)
TDSDDVNKSDIPSFLR
SDIPSFLR / 32
83
53
56
32
53
145
85
-
91
50
130
35 / 110
25 / Hypothetical protein FTL_0637 (gi|89256028) / 532 / 37 / 25.55/5.15 / 25.88/5.4 / DTYTYTYILACEQTR (C)
QAVIIDSVR
FNVNQYLK
LIYAIDTHVHADHVTAAGILR
KVFDGDILEFGNYQLK
VFDGDILEFGNYQLK
LPPPNYIDIAVPANLK / 92
19
54
109
53
95
71 / 126
26 / Dihydrolipoamide dehydrogenase (gi|89255742) / 606 / 22 / 50.50/5.71 / 54.59/6.55 / TQVVVLGSGPGGYSAAFR
AADLGLEVVLVER
YAEIGGVCLNVGCIPSK (2C)
HLESLGIIEMGGLK (M)
IAFYNCIIAAGSSVIK (C)
LPFVPEDDR
GVFPWAASGR
ILFDENHK / 108
92
60
71
104
67
55
50 / 125
27 / Acetyl-CoA acetyltransferase (gi|167009556) / 770 / 45 / 41.70/5.88 / 49.11/6.6 / KRPDELLADVLR
RPDELLADVLR
TVAEANINPNDIEDIVVGCAMPEAEQGLNVAR (C, M)
ISSLLAGLPNSVPAFTINR
QDQDAFALESHQK
AIHAIESGYFK
DEILPIDVDHNKPDEATGEIINHK
LKPAFANGGSVTAGNSSQVSDGAAAVILVSEK (NQ)
FLGFAVAGVDPR
VNPCGGAIALGHPLGATGTILTVK (C) / 56
56
52
87
109
72
48
104
63
124 / 102
28 / Acetyl-CoA acetyltransferase (gi|167009556) / 432 / 23 / 41.70/5.88 / 52.12/6.3 / KRPDELLADVLR
RPDELLADVLR
ISSLLAGLPNSVPAFTINR
QDQDAFALESHQK
AIHAIESGYFK
FLGFAVAGVDPR
VNPCGGAIALGHPLGATGTILTVK (C) / 30
59
100
108
39
38
60 / 128
29 / Inosine-5′-monophosphate dehydrogenase (gi|89256767) / 824 / 39 / 52.06/7.19 / 57.55/7.5 / ITQQAITFDDVLLSPR
YSNVLPHQVDLK
DIQLNIPLVSAAMDTVTESR (M)
LAIAIAQEGGIGIIHK
QESSIKEIMQLAK (NQ)
EHNFSGFPVVDDNNK
DLDEPVSSIMTPR (M)
DNYPHIQVIGGNIATAEAAK
IVAGVGVPQITAIANVAEALK
GTGVPVIADGGIR
KFVPEGVEGR
FVPEGVEGR
TEPTFVQITGAGFNESHVHNVTITK / 117
66
34
49
-
76
5
119
104
45
33
42
64 / 137
30 / Elongation factor EF-Ts (gi|89255659) / 832 / 43 / 30.94/5.57 / 35.07/6.15 / VAAEGVIEVYAADGR
AILLEINSETDFVAR
IGENIQVR
TVEAETLGAYIHGSK
IGVVAALEGGDEDLAK
DVAMHVAAANPMVVSGDQVPADVVAK (2M)
EKEIFTAQAK
KFLDEVALLGQDFVK
FLDEVALLGQDFVK
FLDEVALLGQDFVKDPAIK / 102
113
45
116
100
62
39
35
102
119 / 140
31 / groEL
(gi|1770287) / 1040 / 34 / 57.37/4.99 / 60.67/5.4 / QVLFSDEAR
EIELEDKFENMGAQIVK (M)
TADVAGDGTTTATVLAQALLTEGLK
EGVITVEEGKGFEDELDVVEGMQFDR (M)
GFEDELDVVEGMQFDR
GFEDELDVVEGMQFDR (M)
DLLPILEGVSK
ALLIIAEDVESEALATLVVNNMR (M)
ANIAEANSDYDREK
AAVEEGIVAGGGVALIR
ALDGLTGENDDQNHGIALLR
KAIEAPLR
QIVSNAGGESSVVVNQVK / 66
47
191
26
107
65
78
16
68
137
97
25
121 / 137
32 / UDP-glucose/GDP mannose dehydrogenase (gi|89255994) / 335 / 16 / 48.83/5.29 / 53.87/5.8 / FHIVAVPTPVK
SGEDFKVGYSPER
VGYSPER
VYELVVDAGVYR
AAELGYYSQVILSGR
VAIFGFTFK
SEFEKDYIYWR
DYIYWR / 51
-
18
63
101
40
20
15 / 133
33 / Peroxidase/catalase (gi|290954552) / 653 / 29 / 59.21/5.52 / 76.44/5.6 / YTQEFYNNPEEFKEEFAK
QVSTQDIAQLEQDIINSGLTNQQLIK
TAWDSASTYR
VSLADLIVLGGNVGVEQAAK
QAGYNIQMPFVPGR (M)
SDGFINYTDGSISADKLPQTLVEK
ASMLDLNIPEMTVLVGGMR (2M)
ASMLDLNIPEMTVLVGGMR (3M)
DGEYIGIDR
WTASPVDLIFGSNSELK / 92
140
34
67
21
125
-
19
39
80 / 129
34 / Elongation factor G (gi|89255668) / 449 / 14 / 77.68/4.97 / 81.13/5.3 / ILFYTGLSHK
INIIDTPGHVDFTIEVER
YLEGGELSEDEIHQGLR
LATDPFVGNLTFIR
VEANVGNPQVAYR
GQYGHVFVR
VEVVTPEDYLGDVMGDLNR (M) / 72
121
69
76
63
68
32 / 133
35 / Dihydrolipoamide acetyltransferase (gi|89255741) / 228 / 15 / 56.76/5.08 / 82.75/5.55 / FGEIETQPLSR
IPHVTFYDDADVTDLEEFR
ITPLSFLVK
AAAVALQEFPR
AAAVALQEFPRFNSSLSNDGENLIIK (2NQ)
DIMELAGK
TAVKPIWNGK (NQ) / 55
129
34
45
-
-
- / 240
36 / Molecular chaperone DnaK (gi|56708329) / 1310 / 37 / 69.21/4.88 / 74.94/5.2 / VIENAEGHR
TTPSVVAYTDSGEILVGQAAK
QAVTNPDNTFFAIK
QAVTNPDNTFFAIKR
MAPPQVSAEVLR (M)
TAEDYLGEPVTEAVITVPAYFNDSQR
RIINEPTAAALAYGVDSK
IINEPTAAALAYGVDSK
EQGIDLHNDKLALQR
AKFESLVSDLVMR (M)
FESLVSDLVMR
SDITEVLLVGGQTR
KDVNPDEAVAVGAAIQGGVLAGDVK
SQVFSTAEDNQPAVTIHVLQGER
FDLADIPPAPR
FHDLVTAR
ALEEAFAPIAQK
ALEEAFAPIAQKAYAEQAQAAVAQGGAK (4NQ) / 35
157
67
34
32
153
9
110
65
25
21
95
126
62
69
60
68
- / 135
37 / Fumerate hydratase (gi|89255932) / 248 / 20 / 55.03/5.27 / 61.47/6.0 / DFIDAMYEAYQR
DFIDAMYEAYQR (M)
MSAIGKRPICQDTGMVCAFVK (C, NQ)
TITELINEGVR
RGYNDPYNPLR
GYNDPYNPLR
ASMVFPPHGSR (M)
NGPQNETEQLR
HVHFVLDGSGAVDLPAPK
AGIMGMIGK / 19
21
-
54
9
29
-
26
73
- / 225
38 / ClpB protein
(gi|89255534) / 115 / 47 / 95.99/5.38 / 76.95/5.95 / DLIATLHKMQGLANK
FRGDFEER
VLVDEPTVEDTIAILR
AIDLVDEAASQIR
ALAEFLFDDEDAMLR / -
7
25
15
8 / 65

a NCBI accession number of identified proteins is mentioned.

bThe sequence coverage of protein, as a percentage

c Theoritacal molecular weight and pI

d Calculated molecular weight and pI

Variable modifications: Carbamidomethyl (C),Deamidated (NQ),Oxidation (M)